1. Field of the Invention
The present invention relates to a gel casting system for preparing gels for electrophoresis.
2. Related Art
A gel for use in electrophoresis is typically cast in a gel mold. A gel mold generally consists of two glass or plastic plates separated by spacers and clamped together along each side. The gel is then inserted in its liquid state between the plates, and a comb is inserted in the gel. The gel is allowed to polymerize, and the comb is then removed. The teeth from the comb create wells in the gel. A sample consisting of a specimen containing DNA, RNA or protein is placed in each well for testing. Once the samples are inserted in the wells, electrophoresis of the gel is performed. The results from the electrophoresis appear directly below each sample in the form of bands. The bands typically extend from the top of the gel below each sample to the bottom edge of the gel. Thus, it is important that the gel cover the entire available surface of the plates of the gel mold to maximize the amount of space for obtaining readable, useful information on each gel. It is also important that the gel be free of defects. The bands on the gel are interpreted by comparing one column of bands under a sample relative to another column of bands on the gel. A defect in the gel may distort the bands at certain points, causing the rows of bands to bend or bow, thus making it difficult for the user to interpret the data.
The gel mold is often filled while positioned vertically. This is often referred to as vertical gel casting. When gel, in its liquid state, is inserted between the glass plates of the gel mold, the gel has a tendency to leak out of the sides and bottom of the gel mold. There are several conventional methods, which are described below, used to prevent leakage when casting vertical gels. However, all of these methods are labor intensive and/or often do not completely solve the problem of leakage.
One common method used to prevent leakage in vertical gel casting is to tape the side and bottom edges of the glass plates of the gel mold. Two glass plates with spacers between them along two sides are taped together. The spacers hold the glass plates apart by a predefined distance. Gel is inserted in the region between the plates and allowed to polymerize. Once the gel has polymerized, the tape is removed, and electrophoresis is performed. The taping method has several drawbacks. The most obvious drawback with this method is that applying tape to the glass plates is time consuming. The user must apply new tape for each gel cast. Another drawback is that even with careful application of the tape, the gel often leaks. It is particularly difficult to apply tape around the two bottom corners of the glass. A fold akin to hospital corners must be used at these corners to prevent leakage. Another drawback is that tape oftentimes leaves a residue on the glass plates. Because the glass plates are reused for subsequent gel casting, this residue must be removed before a new gel is cast using these plates. The extra cleaning to remove residue is time consuming, and any residue left on the plates may cause defects on the new gel. Thus, several other methods for gel casting have been developed to address certain of these problems.
Another conventional device for gel casting consists of a gel mold made from two plates with spacers therebetween, and a binder clip on each side to clamp the plates together. Pressure from the binder clips forms a seal down the sides of the plates to prevent leakage. The space along the bottom edge of the plates is left open. A plug, made from a material such as agarose, is used to seal the bottom edge. The plug is made by pouring a strip of molten agarose on a surface, such as a piece of wax paper. The bottom edge of the gel mold is then pressed onto the strip of agarose, and capillary action between the plates causes the agarose to rise and fill in along the bottom edge of the gel mold. Once the agarose solidifies, a plug is formed. The gel to be used for electrophoresis is then inserted between the plates from the top of the gel mold, and allowed to polymerize. After the gel mold has polymerized, the mold is separated from the plug.
The plug method also has several drawbacks. A gel mold with this type of plug has a tendency to leak if the agarose, or other plug material, does not completely cover the entire bottom surface of the gel mold when the plug is being formed. Leakage is particularly noticeable at the corners of the gel mold. Also, the capillary action between the plates may cause the plug material to rise too far inside the gel mold which causes several problems.
For example, if the plug is removed from the gel mold after the gel has polymerized, a large air pocket along the bottom edge of the gel mold is formed. A gel or a buffer solution must completely cover from the top to the bottom surface of the plates to complete the electrical path required to perform electrophoresis. Thus, the user must fill the air pocket before conducting tests on the gel. Often, this air pocket is filled with a buffer solution, such as an electrolyte, capable of carrying an electrical signal through the gel. Alternately, the user may leave the plug in the bottom of the gel mold. In either case, the gel is effectively shortened by the height of the plug or the space, because the buffer solution or the plug material may possess different qualities than the gel used for electrophoresis. Therefore, data may not be readable at the top of the gel due to poor resolution or at the bottom of the gel because the bands have run into the plug material or the buffer solution, and the sample may have to be tested again to obtain the necessary results.
Another conventional means for preventing leakage during gel casting is by using a casting stand, similar to the one made by Bio-Rad Laboratories, Hercules, Calif., Cat. #165-2943. This apparatus comprises two glass plates separated by spacers, in which the glass plates are secured together with knurled thumb screws. This assembly is then inserted onto a second apparatus which presses the bottom edges of the glass plates firmly against a silicone pad to seal the bottom of the gel mold. The Biorad device also has several drawbacks. Over time, the silicone pad loses its resiliency, and the pressure exerted by the glass plates on the silicone pad creates an indentation on the pad. The indentation causes the pad to lose its sealing qualities, and leakage occurs. Leakage may also occur along the bottom edges of the glass plates if the plates are cut unevenly. This is because the apparatus uses a downward force to seal the gap between the glass plates by pressing the bottom edges against the silicone pad. If the bottom edges of the glass plates are cut unevenly, the downward force will not completely seal bottom edges of the glass plates against the silicone pad, and leakage occurs. The present invention does not encounter this problem because it uses inward pressure on the edges of each outer surface of the glass plates to seal the glass plates and prevent leakage.
The devices described above all relate to methods for casting a single gel. Often a user will want to cast several gels at the same time. In particular, gradient gels are often cast simultaneously. Gradient gels are formed by mixing two different gels with different properties (i.e., one is denser than the other, or one has a different pore size than the other). These gels are typically combined at a linear rate and then inserted into a gel mold. By varying the density of the gels, the user can control the individual spacing between the bands on the gel. This allows the user to fit more readable information on each gel. The gradients produced from each mixture may vary. Thus, to ensure that each gel is cast with the same gradient, gels are cast simultaneously.
Several conventional devices exist for casting multiple gels and for casting gels in which the materials inside the gel mold form a gradient. One such device includes a gradient former connected via a hose to a multi-casting tank. The multi-casting tank consists of a box, in which the user stacks a plurality of pairs of plates. The tank is then filled through a hole in the bottom with a gradient gel mixture either via a peristaltic pump or a gravity feed from the gradient former. The gel fills the tank and forms a plurality of gradient gels simultaneously as the gel polymerizes. One drawback with this system is that gel polymerizes inside the hose, and becomes difficult to clean. Another drawback is that peristaltic pumps are expensive. If the user relies on gravity to feed the gradient mixture from the gradient former to the tank, then the gradient former must be placed high above the tank. Conventional gradient formers typically use a magnetic stirrer to ensure proper mixing of the two liquids. When the gradient former is placed high above the tank for the gravity feed, the task of ensuring proper stirring becomes inconvenient.
Another drawback with this system is that removal of the individual gels from the multi-casting tank is messy and difficult. In this casting system, as the gel fills the casting tank, it fills between the outer surfaces of the plate pairs. After the gel polymerizes, the plates must be pried apart. Then, excess gel must be cleaned from the outer surface of each gel mold. Often, gels are destroyed in this process. Additionally, the gel is expensive, and the waste of gel material inherent in this multi-casting process is costly.
The most common conventional methods for gel casting have now been outlined. As discussed, none of these methods or devices completely solve the problem of leakage, and none of them are particularly quick or easy to use. The present invention solves the problem of leakage, while making gel casting a relatively quick and efficient process.